The glycopeptide antibiotics are among the most important class of drugs used in the treatment of resistant bacterial infections. [(a) Cooper et al., In Vancomycin, A Comprehensive Review of 30 Years of Clinical Experience, 1986; pp 1-5, Park Row Publications, Indianapolis, Ind.; (b) Glycopeptide Antibiotics; Nagarajan, R., Ed.; Marcel Dekker: New York, 1994; (c) Kahne et al., Chem. Rev. 2005, 105, 425.] Vancomycin [McCormick et al., Antibiot. Annu. 1955-1956, 606], teicoplanin [Parenti et al., J.
Antibiot. 1978, 31, 276] and a set of recently approved semisynthetic derivatives, including oritavancin (August 2014) [(a) Nicas et al., Antimicrob. Agents Chemother. 1996, 40, 2194; (b) Nagarajan et al., J. Antibiot. 1989, 42, 63; (c) Markham, Drugs 2014, 74, 1823], dalbavancin (May 2014) [(a) Candiani net al., J. Antimicrob. Chemother. 1999, 44, 179; (b) Anderson et al., Drugs 2008, 68, 639] and telavancin (September 2009) [(a) Judice et al., Bioorg. Med. Chem. Lett. 2003, 13, 4165; (b) Corey et al., Nat. Rev. Drug Discovery 2009, 8, 929] are widely or increasingly used to treat clinically refractory and resistant bacterial infections.
Vancomycin, Compound 1, is the central member of the glycopeptide antibiotics that are among
the most important class of drugs used in the treatment of resistant bacterial infections. [(a) Glycopeptide Antibiotics; Nagarajan, R., Ed.; Marcel Dekker: New York, 1994; (b) Kahne et al., Chem. Rev. 2005, 105, 425.] Although it was disclosed in 1956 [McCormick et al., Antibiot. Annu. 1955-1956, 606], and introduced into the clinic in 1958, the structure of vancomycin was established only 25-30 years later (above). [Harris et al., J. Am. Chem. Soc. 1983, 105, 6915.]
After more than 50 years of clinical use and even with the additional widespread use of glycopeptide antibiotics for agricultural livestock (avoparcin), worldwide observation of vancomycin-resistant pathogens has only slowly emerged. This was first restricted to vancomycin-resistant Enterococci (VRE) initially detected in 1987 after 30 years of clinical use [(a) Leclercq et al., N. Engl. J. Med. 1988, 319, 157; (b) Courvalin, Clin. Infect. Dis. 2006, 42, S25] but recently includes the more feared emergence of vancomycin-resistant Staphylococcus aureus (VRSA) first detected in 2002. [(a) Weigel et al., Science 2003, 302, 1569; (b) Howden et al., Clin. Microbiol. Rev. 2010, 23, 99; (c) Walsh et al., Ann. Rev. Microbiol. 2002, 56, 657]. In spite of the increasing prevalence of VRE, such infections presently remain sensitive to other common antibiotic classes although a time may come when this will no longer be the case.
More significant is the emergence of VRSA, which has already acquired resistance to other common classes of antibiotics. Treatment options in such cases are expected to be limited and, outside the new generation glycopeptide antibiotics, these presently include antibiotics known to easily evoke resistance (linezolide, daptomycin). [(a) Brickner, Curr. Pharm. Des. 1996, 2, 175; (b) Scheetz et al., Antimicrob. Agents Chemother. 2008, 52, 2256; (c) Brickner et al., J. Med. Chem. 2008, 51, 1981; and (a) Baltz et al., Nat. Prod. Rep. 2005, 22, 717; (b) Baltz, Curr. Opin. Chem. Biol. 2009, 13, 144] have been designated or recommended for use as “reserve antibiotics”; ones that should be employed sparingly to preserve their effectiveness as drugs of last resort against intractable infections. This has intensified interest in the development of alternative treatments for resistant pathogens that display the remarkable clinical durability of vancomycin [(a) Cooper et al., In Vancomycin, A Comprehensive Review of 30 Years of Clinical Experience, 1986; pp 1-5, Park Row Publications, Indianapolis, Ind.; (b) Glycopeptide Antibiotics; Nagarajan, R., Ed.; Marcel Dekker: New York, 1994; (c) Kahne et al., Chem. Rev. 2005, 105, 425; (d) Malabarba et al., Med. Res. Rev. 1997, 17, 69; (e) Najarajan et al., Drugs 2004, 64, 913; (f) Butler et al., J. Antibiot. 2014, 67, 631].
Clinical uses of vancomycin include the treatment of patients on dialysis, allergic to β-lactam antibiotics, or undergoing cancer chemotherapy. [(a) Cooper et al., In Vancomycin, A Comprehensive Review of 30 Years of Clinical Experience, 1986; pp 1-5, Park Row Publications, Indianapolis, Ind. (b) Glycopeptide Antibiotics; Nagarajan, R., Ed.; Marcel Dekker: New York, 1994. (c) Kahne et al., Chem. Rev. 2005, 105, 425.] However, the most widely recognized use of vancomycin is the treatment of methicillin-resistant Staphylococcus aureus (MRSA) infections. [(a) Cooper et al., In Vancomycin, A Comprehensive Review of 30 Years of Clinical Experience, 1986; pp 1-5, Park Row Publications, Indianapolis, Ind. (b) Glycopeptide Antibiotics; Nagarajan, R., Ed.; Marcel Dekker: New York, 1994. (c) Kahne et al., Chem. Rev. 2005, 105, 425.] The prevalence of MRSA in intensive care units (ICU, 60% of SA infections in the US are MRSA) [(a) CDC (2003); National Nosocomial Infections Surveillance (NNIS) System Report, Data Summary from January 1992 Through June 2004, Issued October 2004. Am. J. Infect. Control 2004, 32, 470; (b) Laxminarayan, Antibiotic Resistance: The Unfolding Crisis. In Extending the Cure, Policy Responses to the Growing Treat of Antibiotic Resistance, Laxminarayan et al., Eds.; Resources for the Future, 2007, Chapter 1, pp 25-37; (c) Walsh et al., Sci. Am. 2009, 301 (1), 44] and its movement from a hospital-acquired to a community-acquired infection in the last 10 years has increased the number and intensified the need to treat such resistant bacterial infections.
In addition, vancomycin-resistant bacterial strains are also on the rise with US ICU clinical isolates of vancomycin-resistant Enterococcus faecalis (VRE) approaching 30% [(a) CDC (2003); National Nosocomial Infections Surveillance (NNIS) System Report, Data Summary from January 1992 Through June 2004, Issued October 2004. Am. J. Infect. Control 2004, 32, 470; (b) Laxminarayan, Antibiotic Resistance: The Unfolding Crisis. In Extending the Cure, Policy Responses to the Growing Treat of Antibiotic Resistance, Laxminarayan et al., Eds.; Resources for the Future, 2007, Chapter 1, pp 25-37; (c) Walsh et al., Sci. Am. 2009, 301 (1), 44], albeit in strains presently sensitive to other antibiotics. Most feared is the recent emergence of MRSA strains now resistant or insensitive to vancomycin (VRSA and VISA). This poses a major health problem and has intensified efforts to develop antibiotics to not only combat this resistance, but that also display the durability of vancomycin [(a) Harris et al., J. Am. Chem. Soc. 1983, 105, 6915; (b) Williamson et al., J. Am. Chem. Soc. 1981, 103, 6580].
Vancomycin is structurally based on a heptapeptide scaffold that has undergone extensive oxidative cross-linking. Five of the seven residues are aromatic, and each residue is assigned a number in the sequence, beginning with leucine at position 1, and a hydroxyphenylglycine (HPG) at residue position 4.
As is seen from the structural formula above, vancomycin contains two amine groups, a carboxylic acid and three potentially acidic phenolic hydroxyl groups. Vancomycin is reported to have the following pKa values: 7.75, 8.89 (amines; basic), 2.18 (carboxyl), 9.59, 10.4 and 12 (phenolic; acidic) [Vijan, Rev. Roum. Chim. 2009, 54(10), 807-813]. Vancomycin hydrochloride is sold for both oral and parenteral administration.
After more than 50 years of clinical use and with the even more widespread utilization of glycopeptide antibiotics for agricultural livestock (avoparcin), worldwide observation of vancomycin-resistant pathogens has slowly emerged. This was first restricted to vancomycin-resistant Enterococci (VRE) [(a) Leclercq et al., N. Engl. J. Med. 1988, 319, 157; (b) Courvalin, Clin. Infect. Dis. 2006, 42, S25], but more recently includes the detection of vancomycin-resistant Staphylococcus aureus (VRSA) [(a) Weigel et al., Science 2003, 302, 1569; (b) Walsh et al., Ann. Rev. Microbiol. 2002, 56, 657]. Interest has consequently intensified in the development of alternative treatments for resistant pathogens that display the remarkable durability of vancomycin, including new derivatives of the glycopeptide antibiotics [(a) Glycopeptide Antibiotics; Nagarajan, R., Ed.; Marcel Dekker: New York, 1994; (b) Kahne et al., Chem. Rev. 2005, 105, 425; (c) Malabarba et al., Med. Res. Rev. 1997, 17, 69; (d) Najarajan et al., Drugs 2004, 64, 913; (e) Süssmuth, ChemBioChem 2002, 3, 295; (f) et al., Chem. Rev. 2005, 105, 449; (g) von Nussbaum et al., Angew. Chem., Int. Ed. 2006, 45, 5072].
The clinical durability can be attributed to several complementary features of vancomycin that result in inhibition of bacterial cell wall biosynthesis and its integrity. [James et al., ACS Chem. Biol. 2012, 7, 797] Foremost of the features responsible for this durability is its primary biological target (binding to D-Ala-D-Ala). This target is not only unique to bacteria, but it is also a structural component of the bacterial cell wall and a substrate for an enzymatic reaction. It is not a protein or nucleic acid target and, as a consequence, it is not subject to alteration by genetic mutation. Moreover, the ramifications of additional candidate binding sites within the bacterial cell wall (not only D-Ala-D-Ala, but also D-Ala-Gly and Gly-Gly) have yet to be defined.
Vancomycin's primary mechanism of action involves substrate sequestration (D-Ala-D-Ala) for a critical late-stage enzyme (transpeptidase) catalyzed reaction needed for peptidoglycan cross-linking and bacterial cell wall maturation. However, it is thought to also inhibit transglycosylase-catalyzed incorporation of lipid intermediate II into the repeating polysaccharide backbone of the bacterial cell wall. With this second mechanism of action for vancomycin, it is not yet established whether this involves direct binding of the appended disaccharide to the enzyme active site, or whether additional cell wall binding sites (e.g., D-Ala-D-Ala, D-Ala-Gly, or Gly-Gly) contribute to its localization and indirect enzyme inhibition. Because there may be two or more mechanisms of action that contribute to the inhibition of bacterial cell wall maturation by vancomycin, full bacterial resistance may require statistically unlikely simultaneous changes to each to overcome all contributing mechanisms.
Just as importantly, the site of action is at the bacterial cell wall surface and not at an intracellular target. As a result, no bacterial cell wall penetration or import mechanism is needed and this permits vancomycin to avoid the common resistance mechanisms mediated by efflux pumps, blocked transport, and deactivation by cytosolic metabolic enzymes. [(a) Wright, Chem. Commun. 2011, 47, 4055; (b) Walsh, C. T. Nature, 2000, 406, 775]
Regardless of the origin and it is likely there are additional features contributing to the durability of vancomycin that are not yet recognized, it is most revealing that the primary mechanism of resistance to the glycopeptide antibiotics (VanA and VanB) was transferred to pathogenic bacteria from non-pathogenic producing organisms that use this inducible mechanism to protect themselves during vancomycin production. [Marshall et al., Antimicrob. Agents Chemother. 1998, 42, 2215] Significantly, this highlights that pathogenic bacteria have not yet independently evolved effective resistance mechanisms to the glycopeptide antibiotics even after more than 50 years of widespread use [identified mechanisms of resistance: VanA and VanB (inducible D-Ala-D-Ala to D-Ala-D-Lac, 1000-fold), VanC (D-Ala-D-Ser, 20-fold), and thickened cell wall (increased number of target sites, 10-fold). See: Courvalin, Clin. Infect. Dis. 2006, 42, S25], suggesting that fundamental solutions to VanA and VanB resistance may provide durable antibiotics with clinical lifetimes lasting 50 more years.
Due to their structural complexity, essentially all analogues of the glycopeptide antibiotics consist of semisynthetic derivatives of the natural products. [(a) Cooper et al., In Vancomycin, A Comprehensive Review of 30 Years of Clinical Experience, 1986; pp 1-5, Park Row Publications, Indianapolis, Ind.; (b) Glycopeptide Antibiotics; Nagarajan, R., Ed.; Marcel Dekker: New York, 1994; (c) Kahne et al., Chem. Rev. 2005, 105, 425; (d) Malabarba, et al., Med. Res. Rev. 1997, 17, 69; (e) Najarajan, J. Antibiot. 1993, 46, 1181; (f) Van Bambeke et al., Drugs 2004, 64, 913; (g) Butler et al., J. Antibiot. 2014, 67, 631] The most significant of the modifications introduce peripheral hydrophobic groups and these are found in each of the clinically approved semisynthetic derivatives oritavancin, dalbavancin and telavancin, whose structural formulas are shown below.

For both dalbavancin and telavancin, the long chain hydrophobic alkyl chains are thought to provide selective membrane anchoring properties and promote antibiotic dimerization without impacting binding affinity to the primary biological target D-Ala-D-Ala. [a) Allen et al., Antimicrob. Agents Chemother. 1996, 40, 2356; (b) Sharman et al., J. Am. Chem. Soc. 1997, 119, 12041; (c) Allen et al., FEMS Microbiol. Rev. 2003, 26, 511] It is possible such semisynthetic changes to the glycopeptide antibiotics also avoid bacterial sensing of the antibiotic challenge and this may account for their VanB VRE activity first observed with teicoplanin. [(a) Hong et al., Adv. Exp. Med. Biol. 2008, 631, 200; (b) Koteva et al., Nat. Chem. Biol. 2010, 6, 327; (c) Ikeda et al., J. Antibiot. 2010, 63, 533; (d) Kwun et al., Antimicrob. Agents Chemother. 2013, 57, 4470] Additionally, telavancin has been shown to function not only through the traditional mechanism of inhibition of cell wall synthesis by binding D-Ala-D-Ala, but also through the disruption of bacterial membrane integrity, a mechanism typically not observed for the glycopeptide antibiotics. [Higgins et al., Antimicrob. Agents Chemother. 2005, 49, 1127]
One of the most widely recognized modifications is the 4-chlorobiphenyl substitution of a peripheral carbohydrate. This substitution has been examined at range of positions in a variety of glycopeptide antibiotics, most notably in oritavancin [(a) Nicas et al., Antimicrob. Agents Chemother. 1996, 40, 2194; (b) Nagarajan et al., J. Antibiot. 1989, 42, 63. (b) Markham, A. Drugs 2014, 74, 1823], the N-(4-chlorobiphenyl)methyl derivative of chloroeremomycin, and with vancomycin itself (CBP-vancomycin). [Cooper et al., J. Antibiot. 1996, 49, 575]
In addition to promoting antibiotic dimerization, membrane anchoring, disruption of bacterial membrane integrity, and potentially avoiding bacterial sensing of the antibiotic challenge, the unique placement of the 4-chlorobiphenyl substituent introduces or potentiates a second mechanism of action. The direct inhibition of transglycosylases mediated by the modified carbohydrate has been identified as a second, now effective, mechanism by which oritavancin exhibits antimicrobial activity. [(a) Ge et al., Science 1999, 284, 507; (b) Chen et al., Proc. Natl. Acad. Sci. USA 2003, 100, 5658; (c) Goldman et al., Microbiol. Lett. 2000, 183, 209]
Regardless of the origin of the effects, such derivatives often increase antibiotic potency as much as 100-fold. Although increasing bacterial sensitivity to the antibiotics, VanA vancomycin-resistant bacterial strains (MIC=about 10 μg/mL) remain 1000-fold less sensitive than susceptible strains (MIC=about 0.01 μg/mL). This suggested that combining such peripheral hydrophobic substitutions with vancomycin binding pocket modifications that maintain D-Ala-D-Ala binding and reinstate binding to D-Ala-D-Lac would further increase their antimicrobial activity against not only sensitive, but also vancomycin-resistant bacteria to truly remarkable potencies.
Recently, and in an extension of work first directed at the total syntheses of the naturally occurring glycopeptide antibiotics [(a) Boger et al., J. Am. Chem. Soc. 1999, 121, 3226; (b) Boger et al., J. Am. Chem. Soc. 1999, 121, 10004; (c) Boger et al., J. Am. Chem. Soc. 2000, 122, 7416; (d) Boger et al., J. Am. Chem. Soc. 2001, 123, 1862; (e) Crowley et al., J. Am. Chem. Soc. 2004, 126, 4310; (f) Garfunkle et al., J. Am. Chem. Soc. 2009, 131, 16036; (g) Shimamura et al., J. Am. Chem. Soc. 2010, 132, 7776; (h) Breazzano et al., J. Am. Chem. Soc. 2011, 133, 18495; (i) James et al., ACS Chem. Biol. 2012, 7, 797; (j) Boger, Med. Res. Rev. 2001, 21, 356; (k) Evans et al., Angew. Chem., Int. Ed. 1998, 37, 2700; (l) Evans et al., Angew. Chem., Int. Ed. 1998, 37, 2704; (m) Evans et al., J. Am. Chem. Soc. 1997, 119, 3419; (n) Evans et al., J. Am. Chem. Soc. 1997, 119, 3417; (o) Nicolaou et al., Angew. Chem., Int. Ed. 1998, 37, 2717; (p) Nicolaou et al., M. Angew. Chem., Int. Ed. 1998, 37, 2708; (q) Nicolaou et al., Angew. Chem., Int. Ed. 1998, 37, 2714; (r) Boger, Med. Res. Rev. 2001, 21, 356; (s) Nicolaou et al., Angew. Chem., Int. Ed. 1999, 38, 2096; (t) Evans et al., Drugs Pharm. Sci. 1994, 63, 63] the present inventor and co-workers described studies on the binding pocket redesign of vancomycin [James et al., ACS Chem. Biol. 2012, 7, 797] that are the first to directly address the molecular basis of clinical resistance to vancomycin. [(a) Bugg et al., Biochemistry 1991, 30, 10408; Reviews: (b) Walsh, Science 1993, 261, 308; (c) Walsh et al., Chem. Biol. 1996, 3, 21; (d) Lessard et al., Proc. Natl. Acad. Sci. USA 1999, 96, 11028; (e) Healy et al., Chem. Biol. 2000, 7, R109; (f) Perkins, Pharmacol. Ther. 1982, 16, 181; (g) Williams et al., J. Am. Chem. Soc. 1983, 105, 1332; (h) Schaefer et al., Structure 1996, 4, 1509]
The destabilized binding to D-Ala-D-Lac is due to a combination of the loss of a H-bond central to ligand binding the antibiotic (10-fold), and an even more significant destabilizing lone pair repulsion between the vancomycin residue 4 carbonyl and D-Ala-D-Lac ester oxygens (100-fold). [McComas et al., J. Am. Chem. Soc. 2003, 125, 9314] The elucidation of this inducible mechanism of resistance (VanA and VanB) acquired from non-pathogenic vancomycin-producing organisms [Marshall et al., Antimicrob. Agents Chemother. 1998, 42, 2215] also highlighted that such vancomycin binding pocket modifications must target compounds that not only establish binding to D-Ala-D-Lac, but that also maintain D-Ala-D-Ala binding. That targeting not only insures antimicrobial activity against vancomycin-resistant bacteria (VanA and VanB), but additionally assures maintained activity against vancomycin-sensitive bacteria.
Previous studies of the inventor and co-workers provided [Ψ[CH2NH]Tpg4]vancomycin aglycon (Compound 10) [Crowley et al., Am. Chem. Soc. 2006, 128, 2885], which displayed such dual binding properties by virtue of removal of the lone pair repulsion between the vancomycin residue 4 carbonyl and D-Ala-D-Lac ester oxygens. This change reinstated commensurate activity against VanA VRE, validated the opportunities of the approach, and entailed removal of a single atom from the vancomycin binding pocket.
These efforts were followed by the total synthesis of [Ψ[C(═NH)NH]Tpg4]vancomycin aglycon (Compound 9) [(a) Xie et al., J. Am. Chem. Soc. 2011, 133, 13946; and (b) Xie et al., J. Am. Chem. Soc. 2012, 134, 1284], providing a modified antibiotic that not only maintained vancomycin's ability to bind the unaltered peptidoglycan D-Ala-D-Ala, but that also bound the altered ligand D-Ala-D-Lac just as effectively by virtue of its ability to serve as either a H-bond donor (for D-Ala-D-Lac) or H-bond acceptor (for D-Ala-D-Ala). Whereas the former entails binding of the protonated amidine with D-Ala-D-Lac and replaces the destabilizing carbonyl lone pair interaction with the ester oxygen lone pair with a stabilizing electrostatic interaction and perhaps a reversed H-bond, the latter entails binding of D-Ala-D-Ala with the unprotonated amidine serving as a H-bond acceptor. [Okano et al., J. Am, Chem. Soc. 2012, 134, 8790] Not only did amidine Compound 9 display balanced binding affinity for both target ligands within 2-fold of that which vancomycin aglycon exhibits with D-Ala-D-Ala, but it also exhibited effective antimicrobial activity against VanA VRE, being equipotent to the activity that vancomycin displays against sensitive bacterial strains.
These latter studies represented the replacement of a single atom in the binding pocket of the antibiotic aglycon (O→NH) to counter a complementary exchange in the cell wall precursors of resistant bacteria (NH→O). Just as remarkable, it was established that [Ψ[C(═S)NH]Tpg4]vancomycin aglycon (Compound 8), which served as the penultimate precursor to Compound 9 [(a) Xie et al., J. Am. Chem. Soc. 2011, 133, 13946; (b) Xie et al., J. Am. Chem. Soc. 2012, 134, 1284], fails to bind D-Ala-D-Ala or D-Ala-D-Lac to any appreciable extent and is inactive against both vancomycin-sensitive and vancomycin-resistant bacteria.
The expectedly benign conversion of the residue 4 amide to a thioamide with the exchange of a single atom in the binding pocket (O→S) proved sufficient to completely disrupt ligand binding. This loss in affinity was attributed largely to the increased thiocarbonyl bond length and size of the sulfur atom that are sufficient to sterically displace the ligand out of the binding pocket and completely disrupt the intricate binding of D-Ala-D-Ala. Significantly, the comparison of Compound 8 with Compound 9 highlighted just how remarkable the behavior of the amidine Compound 9 is. These aglycon structures and data are shown below.
 ligand, Ka (M−1)VanAacompound11, Y = NH12, Y = OKa(11/12)MIC, μg/mL 7, X = O1.7 × 1051.2 × 1021400   640 8, X = S1.7 × 1021.1 × 101—>640  9, X = NH7.3 × 1046.9 × 104  1.05   1b10, X = H24.8 × 1035.2 × 103 0.9  31aMinimum inhibitory conc., E. faecalis (BM4166, VanA VRE).bTested herein alongside Compounds 1-6.
The glycopeptide antibiotics inhibit bacterial cell wall synthesis by binding the precursor peptidoglycan peptide terminus D-Ala-D-Ala, inhibiting transpeptidase-catalyzed cell wall cross-linking and maturation [(a) Perkins, Pharmacol. Ther. 1982, 16, 181; (b) Williams et al., J. Am. Chem. Soc. 1983, 105, 1332; (c) Schaefer et al., Structure 1996, 4, 1509].
In the clinically prominent resistant phenotypes, VanA and VanB, synthesis of the precursor lipid intermediates I and II continue complete with their pendant N-terminal D-Ala-D-Ala, but resistant bacteria sense the antibiotic challenge [(a) Hong et al., J. Adv. Exp. Med. Biol. 2008, 631, 200; (b) Koteva et al., Nat. Chem. Biol. 2010, 6, 327; (c) Ikeda et al., J. Antibiot. 2010, 63, 533; (d) Kwun et al., Antimicrob. Agents Chemother. 2013, 57, 4470] and initiate a late stage remodeling of their peptidoglycan termini from D-Ala-D-Ala to D-Ala-D-Lac [(a) Bugg et al., Biochemistry 1991, 30, 10408; (b) Walsh, Science 1993, 261, 308] to avoid the antibiotic action.
Through use of a two-component cell surface receptor sensing and subsequent intracellular signaling system [(a) Hong et al., Adv. Exp. Med. Biol. 2008, 631, 200; (b) Koteva et al., Nat. Chem. Biol. 2010, 6, 327; (c) Ikeda et al., J. Antibiot. 2010, 63, 533; (d) Kwun et al., Antimicrob. Agents Chemother. 2013, 57, 4470], resistant bacteria initiate a late stage remodeling of their peptidoglycan termini from D-Ala-D-Ala to D-Ala-D-Lac [(a) Bugg et al., Biochemistry 1991, 30, 10408; Reviews: (b) Walsh, Science 1993, 261, 308; (c) Walsh et al., Chem. Biol. 1996, 3, 21; (d) Lessard et al., Proc. Natl. Acad. Sci. USA 1999, 96, 11028; (e) Healy et al., Chem. Biol. 2000, 7, R109; (f) Perkins, Pharmacol. Ther. 1982, 16, 181; (g) Williams et al., J. Am. Chem. Soc. 1983, 105, 1332; (h) Schaefer et al., Structure 1996, 4, 1509] to avoid the action of the antibiotic. The vancomycin binding affinity for this altered ligand is reduced 1000-fold [(a) Bugg et al., Biochemistry 1991, 30, 10408; Reviews: (b) Walsh, Science 1993, 261, 308; (c) Walsh et al., Chem. Biol. 1996, 3, 21; (d) Lessard et al., Proc. Natl. Acad. Sci. USA 1999, 96, 11028; (e) Healy et al., Chem. Biol. 2000, 7, R109; (f) Perkins, Pharmacol. Ther. 1982, 16, 181; (g) Williams et al., J. Am. Chem. Soc. 1983, 105, 1332; (h) Schaefer et al., Structure 1996, 4, 1509] resulting in a corresponding 1000-fold loss in antimicrobial activity.
The direct inhibition of transglycosylases mediated by a glycopeptide-modified carbohydrate has been implicated as a second mechanism by which the lipophilic glycopeptides with impaired D-Ala-D-Lac or D-Ala-D-Ala binding properties exhibit antimicrobial effects [(a) Ge et al., Science 1999, 284, 507; (b) Chen et al., Proc. Natl. Acad. Sci. USA 2003, 100, 5658]. Other compounds, including telavancin, have been shown to function both through the traditional mechanism of inhibition of cell wall synthesis by binding to D-Ala-D-Ala and also through the disruption of bacterial membrane integrity, a mechanism typically not observed for glycopeptides [(a) Higgins et al., Antimicrob. Agents Chemother. 2005, 49, 1127; (b) Corey et al., Nat. Rev. Drug Discovery 2009, 8, 929].
Regardless of the origin of the effect, such derivatives typically increase antibiotic potency as much as 100-fold. While increasing bacterial sensitivity to the antibiotics, VanA vancomycin-resistant bacterial strains (MIC=about 10 μg/mL) remain 1000-fold less sensitive than susceptible strains (MIC=about 0.01 μg/mL).
Because of their structural complexity, essentially all new analogs of the glycopeptide antibiotics consist of semisynthetic derivatives of the natural products [(a) Glycopeptide Antibiotics; Nagarajan, R., Ed.; Marcel Dekker: New York, 1994; (b) Kahne et al., Chem. Rev. 2005, 105, 425; (c) Malabarba et al., Med. Res. Rev. 1997, 17, 69; (d) Najarajan et al., Drugs 2004, 64, 913]. The most significant of the modifications introduce peripheral hydrophobic groups into the glycopeptide structure.
Among the early-reported successes were those of Nagarajan et al., J. Antibiot. 1989, 42, 63 and Nagarajan, R. J. Antibiot. 1993, 46, 118 who disclosed N-decyl, N-p-octylbenzyl and N-p-octyl-oxybenzyl groups bonded to the 4-epi-vancosaminyl substituent of a glycopeptide antibiotic provided enhanced potency against both sensitive and resistant Enterococci. See also, U.S. Pat. Nos. 5,840,684 and 5,843,889.
One later-developed, hydrophobe-modified glycopeptide is telavancin [Corey et al., Nat. Rev. Drug Discovery 2009, 8, 929-930; formerly referred to as TD-6424] a clinically approved (2009) semisynthetic derivative of vancomycin used to treat complicated skin infections that are suspected or confirmed to be MRSA. This drug bears a hydrophobic N-ethylene-2-amino-N-decyl group bonded to the vancosaminyl nitrogen to grant increased activity against resistant organisms and a hydrophilic phosphonic acid side chain that provides improved pharmacokinetic properties. Oritavancin is another widely studied hydrophobically-substituted vancomycin-like glycopeptide derivative that contains a N-4-(4′-chlorobiphenyl)methyl group bonded to the amino nitrogen of a L-4-epi-vancosaminyl-1,2-D-glucoside [(a) Malabarba et al., Med. Res. Rev. 1997, 17, 69; (b) Najarajan et al., Drugs 2004, 64, 913], and a second L-4-epi-vancosaminyl substituent bonded to the cyclic core.
Dalbavancin and teicoplanin are other hydrophobe-substituted glycopeptide antibiotics. Teicoplanin is a natural product that contains a heptapeptide cyclic core structure similar to that of vancomycin but contains four internal cross-links as compared to the three cross-links present in vancomycin. Teicoplanin also contains an N-acetyl-β-D-glucosamine and a D-mannose group separately bonded to the cyclic structure, as well as one of at least five different C10-11-acyl-β-D-glucosamine groups. Dalbavancin contains a cyclic core (scaffold) structure slightly different from teicoplanin, as well as a carboxyl-substituted, N—C10-amidohexoside group, a 3-(dimethylaminopropyl)amido group and a D-mannose group that are bonded to the cyclic core, but lacks an N-acetyl-β-D-glucosamine group present in teicoplanin.
These hydrophobic modifications have been explored in a variety of glycopeptide antibiotics and at range of positions, most notably in oritavancin [(a) Nicas et al., Antimicrob. Agents Chemother. 1996, 40, 2194; (b) Nagarajan et al., J. Antibiot. 1989, 42, 63], the N-(4-chlorobiphenyl)methyl derivative of chloroeremomycin, and with vancomycin itself (Compound 4, CBP-vancomycin) [Kahne et al., Chem. Rev. 2005, 105, 425]. Oritavancin, dalbavancin, teicoplanin, telavancin and similar glycopeptide antibiotics on which these modifications have been tried all have one or more of different glycosyl groups, different side chain substituents, or one or more additional glycosyl groups compared to vancomycin.
Kahne et al., Chem. Rev. 2005, 105, 425 reported that the minimum inhibitory concentration (MIC) against vancomycin-sensitive and -resistant strains of E. faecium of vancomycin itself and CBP-vancomycin were 1 and 2048 μg/mL (vancomycin) vs. 0.03 and 16 μg/mL (CBP-vancomycin). Thus, the activity increased in the presence of the CBP group, but the vancomycin-resistant strain was still about 500-times less sensitive than the sensitive strain.
Studies on the mechanism of action and have shown that the N-4-(4′-chlorobiphenyl)methyl side chain promotes antibiotic dimerization and membrane anchoring and establishes antimicrobial activity against vancomycin-resistant organisms despite a lack of improved binding with either D-Ala-D-Ala or D-Ala-D-Lac [(a) Allen et al., Antimicrob. Agents Chemother. 1996, 40, 2356; (b) Sharman et al., J. Am. Chem. Soc. 1997, 119, 12041; (c) Allen et al., FEMS Microbiol. Rev. 2003, 26, 511]. It is possible such semisynthetic changes to vancomycin also avoid bacterial sensing of the antibiotic challenge and this may account for their VanB VRE activity (like teicoplanin) [(a) Hong et al., J. Adv. Exp. Med. Biol. 2008, 631, 200; (b) Koteva et al., Nat. Chem. Biol. 2010, 6, 327; (c) Ikeda et al., J. Antibiot. 2010, 63, 533; (d) Kwun et al., Antimicrob. Agents Chemother. 2013, 57, 4470], or that they may entail a second mechanism of action.
The full details of the total synthesis of the recently disclosed [Okano et al., J. Am. Chem. Soc. 2014, 136, 13522] [Ψ[C(═NH)NH]Tpg4]vancomycin and [Ψ[C(═S)NH]Tpg4]-vancomycin, and their (4-chloro-biphenyl)methyl derivatives are provided hereinafter. Analogous and previously unreported studies first developed with their corresponding synthetic C-terminal hydroxymethyl precursors, as well as the total synthesis of [Ψ[CH2NH]Tpg4]vancomycin and their corresponding (4-chlorobiphenyl)methyl derivatives are also reported. The latter previously undisclosed studies complete an initial series of totally synthetic vancomycin analogs bearing the peripheral L-vancosaminyl-1,2-D-glucosyl disaccharide as well as their (4-chlorobiphenyl)methyl derivatives.
Collectively, the compounds represent a key set of analogues of vancomycin and its (4-chloro-biphenyl)methyl derivative containing single atom changes in the binding pocket. Their assessments indicate that combined pocket and chiorobiphenyl (CBP) peripherally modified analogues exhibit a remarkable spectrum of antimicrobial activity (VSSA, MRSA, VanA and VanB VRE) and impressive potencies against both vancomycin-sensitive and vancomycin-resistant bacteria, and likely benefit from two independent and synergistic mechanisms of action. Like vancomycin, such analogues are likely to display especially durable antibiotic activity not prone to rapidly acquired clinical resistance.